α2-Macroglobulin (α2M) regulates growth and gene expression in many cell types by binding and neutralizing transforming growth factor β (TGF-β). In this study we characterized the effects of the serine proteinase, plasmin, on the interaction of α2M with TGF-β1 and TGF-β2. Binding of both TGF-β isoforms to purified α2M–plasmin complex was primarily non-covalent and reversible. The binding affinity of α2M for TGF-β1 was increased by plasmin; the Kd values were 320 and 84 nM for native α2M and α2M–plasmin respectively. In contrast the affinity of α2M for TGF-β2 was decreased by plasmin; the Kd values were 14 and 80 nM for native α2M and α2M–plasmin respectively. Thrombin decreased the affinity of α2M for TGF-β2 in a similar manner to plasmin. In assays of DNA synthesis in fetal bovine heart endothelial cells, native α2M neutralized the activity of exogenously added TGF-β2, whereas α2M–plasmin, at equivalent concentrations, had almost no effect. Native α2M and methylamine-modified α2M increased platelet-derived growth factor α-receptor expression in vascular smooth-muscle cells, an activity attributed to the neutralization of autocrine TGF-β activity, whereas α2M–plasmin was less effective at the same concentration. These studies demonstrate that the effects of proteinases on the cytokine-binding and cytokine-neutralizing activities of α2M are cytokine-dependent. By reacting with α2M, proteinases might regulate not only the availability of cytokines in the extracellular spaces but also the composition of the cytokine milieu.

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